The secretion of salt, water, and digestive enzymes by the exocrine pancreas is severely compromised in most patients with cystic fibrosis (CF). The disease results from mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) protein which functions as a cAMP-dependent chloride channel. Recent immunolocalization studies demonstrated that the level of expression of CFTR is high in the epithelial cells that line the small pancreatic ducts. In contrast, the level of expression of CFTR is low in large pancreatic ducts, and is undetectable in acinar cells. The ductal epithelial cells comprise only 5-10% of the volume of the pancreas and are buried within the acinar tissue, and thus poorly accessible for study. Because of these limitations, most conclusions about ductal cell transport function are based upon the effects of transport inhibitors on the volume and composition of secretions collected either in vivo or in vitro from the whole pancreas. Consequently, our knowledge of normal ductal cell transport function is meager, and our understanding of pancreatic pathophysiology in CF is limited. The overall objective of this proposal is to obtain a quantitative understanding of the transepithelial transport properties of small pancreatic ducts, and to determine the precise role of CFTR in ductal sodium bicarbonate secretion. Specifically, we will use isolated perfused intra- and inter-lobular ductal segments to determine the basal and secretin stimulated rates of fluid secretion, the ionic composition of the secreted fluid, and the transepithelial electrical properties of the ductal epithelium. Conventional and ion-selective intracellular microelectrode techniques win be used to identify transport pathways and to determine electrochemical driving forces at the apical and basolateral cell membranes responsible for transepithelial transport. Finally, the extent to which differentiated transport properties are retained by ductal epithelial cells grown in monolayer culture will be determined. The information obtained from these studies will be used to critically evaluate current models of pancreatic ductal secretion, more clearly define the role of CFTR in ductal function, and to provide a basis for evaluation of immortalized ductal epithelial cell lines.